Melanoma remains the deadliest form of skin cancer whose prognosis at five years is still only 15%, underscoring a lack of effective treatments. The present proposal focuses on the targeting of an MT1-MMP/MMP2 signaling axis involved in the growth and progression of melanoma by a novel MT1-MMP/MMP specific inhibitor. MT1-MMP is a membrane associated matrix metalloproteinase that controls pericellular proteolysis and is an important, invasion-promoting, pro-tumorigenic MMP in cancer. Our recently published data show that deregulation deregulation of MT1-MMP expression is an early event and continues to increase during melanoma progression. MT1-MMP expression is associated with poor melanoma patient outcome, underscoring a pivotal role of MT1-MMP in melanoma pathogenesis. Indeed, MT1-MMP is required for melanoma cells to metastasize, as cells deprived of MT1-MMP fail to form distant metastasis in an orthotopic mouse melanoma model. MT1-MMP affects cell invasion by activating its target MMP2, which isalso required by MT1-MMP to sustain RAC1 activity and promote MT1-MMP dependent cell motility, highlighting a novel MT1-MMP/MMP2/RAC1 signaling axis in melanoma. Yet, preliminary data show that while MT1-MMP also controls cell growth, MMP2 does not, pointing to distinct roles of the two proteases in melanoma growth and dissemination. Nevertheless, our data indicate both functions can be therapeutically targeted by the novel MT1-MMP/MMP2 inhibitor ND-322 in several melanoma cell lines. ND-322 diminishes melanoma cell migration, invasion and growth, similarly to specific knock down of MT1-MMP and/or MMP2. Importantly, while the lack of specificity of broad-spectrum MMP inhibitors have been accompanied by severe side effects, such as musculoskeletal pain and inflammation due to the targeting of both the good and the bad MMPs, ND-322 has shown high tolerability in mouse models of brain ischemia. Hence, we have hypothesized that the specific MT1-MMP/MMP2 inhibitor ND-322 represent a safe, novel valid addition to the current treatment options available to patients that is capable of effectivel inhibit melanoma growth and metastasis. Aim 1 will assess the efficacy of ND-322 in association with Vemurafenib (BRAF inhibitor) in a BRAFV600E transgenic mouse melanoma model that develops lesions that genetically and biologically resemble the human disease and that expresses both MT1-MMP and MMP2. The second aim will characterize, in an orthotopic mouse melanoma model the roles of MT1-MMP and MMP2 downstream of ND-322 in modulating melanoma growth and metastasis, by utilizing human melanoma cells expressing specific shRNAs against MT1-MMP or MMP2. The knowledge gained from this work will enhance our understanding of the mechanisms driving melanoma growth and progression while providing the rationale for a treatment approach employing a novel MMP inhibitor.